2019—2022年聊城市食品和病例分离单增李斯特菌基于全基因组测序的分子特征及耐药性分析

Molecular characterization and drug resistance analysis based on whole-genome sequencing of Listeria monocytogenes isolated from food and patients in Liaocheng City from 2019 to 2022

目的基于全基因组测序(WGS)比较分析2019—2022年聊城市单核细胞增生李斯特菌(Lm)食品和病例分离株基因组特征、毒力性、耐药性以及遗传多样性。方法对聊城市33株市售食品和临床病例中Lm分离株开展抗生素敏感性试验和WGS。利用MGAP对WGS数据进行拼接组装,对组装基因组进行基因预测和功能注释、MLST,制作cg MLST最小生成树图,并与美国国家生物信息中心(NCBI)上获取的18株国内外Lm构建wg-SNP进化树。结果33株Lm分离株的基因组大小为2.89~3.41 Mb,CG含量为37.81%~37.97%,可分为6个ST型(ST9、ST121、ST8、ST87、ST155、ST101),分别属于6个克隆复合群(CC9、CC121、CC8、CC87、CC155、CC101);分离株均携带fosX和mprF耐药基因,此外还携带lplA1、prsA2等其他18个毒力基因,有不同程度的毒力基因缺失情况。2株菌对四环素耐药,1株菌对林可霉素耐药。均携带毒力岛LIPI-1和LIPI-2,未检测到毒力岛LIPI-3和LIPI-4。wg-SNPs、cgMLST和基于单拷贝核心蛋白序列的系统发育树遗传进化分析显示,33株Lm分子分型呈现高度多样性,病例来源菌株与食品分离株亲缘关系密切,食品分离株与国外暴发分离株在进化关系上密切相关。结论山东省聊城市食品和病例中分离的单增李斯特菌均携带毒力基因,具有一定的潜在致病能力,耐药情况尚不严重。分子型别呈现出多样性,食品来源菌株和病例分离株具有较近的亲缘关系,提示市售食品有食源性感染的潜在风险。

Objective The genomic characteristics,toxicity,drug resistance and genetic diversity of Listeria monocytogenes(Lm)isolated from food and patients based on whole genome sequencing(WGS)in Liaocheng City from 2019 to 2022 were investigated.Methods The Lm strains isolated from food and patients were detected by the drug sensitivity test and WGS.The whole genome sequencing data was spliced and assembled by MGAP to carry out gene prediction and functional annotation on the assembled genome,MLST,production of cg MLST minimum spanning tree.The genetic evolution of 18 domestic and foreign foodborne Lm isolated strains obtained from NCBI were analyzed by wgSNP method.Results The genome size of 33 Lm isolates was 2.89-3.41 Mb,and the CG content ranged from 37.81%to 37.97%.The strains could be divided into 6 ST types(ST9,ST121,ST8,ST87,ST155,ST101),corresponding to 6clonal complex groups(CC9,CC121,CC8,CC87,CC155,CC101).The strains carried fosX and mprF resistance gene,18 other virulence genes such as lplA1 and prsA2,with varying degrees of deletion of virulence genes.2 strains were resistant to tetracycline and 1 strain was resistant to lincomycin.The stains carried virulence islands LIPI-1 and LIPI-2without virulence islands LIPI-3 and LIPI-4.The genetic evolution analysis of wg-SNPs,cgMLST and phylogenetic tree based on single-copy core protein sequence showed that the molecular typing of 33 Lm strains was with high diversity in molecular typing.Patient derived strains were closely related to food isolates,and food isolates were closely related to foreign outbreak isolates in evolutionary relationships.Conclusion The strains of Lm isolated from food and patients both carry virulence genes with certain potential pathogenic ability.The drug resistance of the isolated strains is not serious.Molecular types exhibit diversity.The close genetic relationship between strains from food sources and patient isolates suggests that the potential risks of foodborne infections in commercially available food.